Molecular analysis of acute rejection on transplanted liver

本研究探讨了L-选择素介导的淋巴细胞归巢机制是否参与了肝移植急性排斥反应的发生。移植后立即活检的肝脏样本以及显示急性排斥反应的肝脏样本用外周淋巴结地址素（PNAd）特异性的MECA-79抗体进行免疫染色。2例PNAd仅在急性排斥反应的汇管区内皮细胞表达。由于L-选择素配体6-磺基唾液酸刘易斯X存在于核心2 O-聚糖的末端，因此使用针对C2 GnT-1的抗血清通过免疫组织化学分析核心2 β 6 GlcNAcT-1（C2 GnT-1）（核心2 O-聚糖生物合成的关键酶）的存在。结果，在PNAd阳性的内皮细胞中未检测到C2 GnT。此外，这些内皮细胞不被对包括6-磺基唾液酸刘易斯X在内的刘易斯X相关碳水化合物具有特异性的HECA-452抗体识别。在与Burnham研究所的Minoru Fukuda教授的合作研究中，通过克隆一种新的糖基转移酶corel extension β1，3GlcNAcT，表明PNAd是Galβ1→4（sulfo→6）GlcNAcβ1→3Galβ1 →3GalNAc。相同的研究还表明，6-磺基唾液酸刘易斯X不仅在核心2 O-聚糖的末端表达，而且在延伸的核心1 O-聚糖的末端表达，并作为L-选择素的配体。综上所述，本研究表明，PNAd可以表达在内皮细胞的急性排斥反应。6-sulfosialyl刘易斯X在PNAd阳性内皮细胞中不表达，提示L-选择素介导的淋巴细胞归巢与移植肝急性排斥反应的发生无关。

We have tested if lymphocyte homing mechanism mediated by L-selectin is involved in the pathogenesis of acute rejection of transplanted liver. Liver samples biopsied just after the transplantation as well as showing the acute rejection were immunostained with MECA-79 antibody specific for peripheral node addressin ( PNAd ). In 2 of 8 patients, PNAd was found to be expressed only in the endothelial cells in portal area exhibiting acute rejection. Since L-selectin ligand, 6-sulfo sialyl Lewis X is present in the terminal end of core2 O-glycans, the presence of core2β6GlcNAcT-l ( C2GnT-l ), a key enzyme for the biosynthesis of core2 O-glycans was analyzed by immunohistochemistry using antisera against C2GnT-l. As results, C2GnT was not detectable in the endothelial cells positive for PNAd. In addition, these endothelial cells were not recognized by HECA-452 antibody specific for Lewis X-related carbohydrates including 6-sulfo sialyl Lewis X. In a collaborative study with Professor Minoru Fukuda at the Burnham Institute, it is shown that PNAd is Galβ1→4( sulfo→6 )GlcNAcβ1→3Galβ1→3GalNAc by cloning a novel glycosyltransferase, corel extension β1, 3GlcNAcT. The same study also demonstrates that 6-sulfo sialyl Lewis X is expressed in the terminal ends of not only core2 O-glycans but also extended corel O-glycans and serves as ligand for L-selectin. Taken together, the present study indicates that PNAd can be expressed in the endothelial cells showing acute rejection. However, 6-sulfo sialyl Lewis X was not expressed in the PNAd-positive endothelial cells, suggesting that the L-selectin-mediated lymphocyte homing is barely involved in the pathogenesis of acute rejection of the transplanted liver.