Holán Vladimír, Pindjáková Jana, Krulová Magdaléna, Neuwirth Ales, Fric Jan, Zajícová Alena
Institute of Molecular Genetics, Academy of Sciences, and Faculty of Natural Sciences, Charles University, Prague, Czech Republic.
Transplantation. 2006 Jun 27;81(12):1708-15. doi: 10.1097/01.tp.0000226067.89690.2b.
Production of nitric oxide (NO) by graft infiltrating macrophages has been proposed as an important effector mechanism of allograft rejection. Although high levels of NO are generated during allograft rejection, undetectable or only limited amounts of NO were found in rejected skin xenografts.
BALB/c mice were grafted with skin transplants from syngeneic, allogeneic or xenogeneic (rat) donors. The production of NO, cytokines and arginase in the grafts was determined by spectrophotometry, enzyme-linked immunosorbent assay, or polymerase chain reaction. Effects of depletion of CD4+ cells, neutralization of interleukin (IL)-4 or application of arginase inhibitors N(omega)-hydroxy-L-arginine (L-NOHA) and L-valine on production of NO in rejected xenografts were evaluated.
Rejection of rat skin xenografts, on the contrary to rejection of allografts, was associated with a local high production of Th2 cytokines IL-4 and IL-10, overexpression of arginase genes, strongly enhanced arginase activity and attenuated NO generation in the graft. The supernatants obtained after cultivation of skin xenograft (but not allograft or syngeneic graft) explants contained a high arginase activity and strongly suppressed NO production by activated macrophages. This suppression was completely inhibited by L-NOHA or was overcome by an excess of exogenous L-arginine, a substrate for NO synthesis. Cocultivation of xenograft explants that did not produce NO with arginase inhibitors L-NOHA or L-valine restored NO generation in the graft.
The results suggest that upregulation of arginase activity by Th2 cytokines during xenograft rejection limits the bioavailability of L-arginine for the inducible NO synthase and thus attenuates generation of NO by the graft-infiltrating macrophages.
移植浸润巨噬细胞产生一氧化氮（NO）被认为是同种异体移植排斥反应的一种重要效应机制。尽管在同种异体移植排斥反应过程中会产生高水平的NO，但在被排斥的皮肤异种移植中却检测不到或仅发现有限量的NO。
将同基因、异基因或异种（大鼠）供体的皮肤移植到BALB/c小鼠身上。通过分光光度法、酶联免疫吸附测定或聚合酶链反应测定移植组织中NO、细胞因子和精氨酸酶的产生。评估CD4+细胞耗竭、白细胞介素（IL）-4中和或应用精氨酸酶抑制剂N（ω）-羟基-L-精氨酸（L-NOHA）和L-缬氨酸对被排斥异种移植组织中NO产生的影响。
与同种异体移植排斥反应相反，大鼠皮肤异种移植排斥反应与局部Th2细胞因子IL-4和IL-10的高产生、精氨酸酶基因的过度表达、精氨酸酶活性的强烈增强以及移植组织中NO生成的减弱有关。皮肤异种移植（而非同种异体移植或同基因移植）外植体培养后获得的上清液含有高精氨酸酶活性，并强烈抑制活化巨噬细胞产生NO。这种抑制作用被L-NOHA完全抑制，或被过量的外源性L-精氨酸（NO合成的底物）克服。用精氨酸酶抑制剂L-NOHA或L-缬氨酸与不产生NO的异种移植外植体共培养可恢复移植组织中NO的生成。
结果表明，异种移植排斥反应期间Th2细胞因子上调精氨酸酶活性，限制了诱导型NO合酶对L-精氨酸的生物利用度，从而减弱了移植浸润巨噬细胞产生NO的能力。
