Ashraf M, Samra Z Q
Department of Pathology and Laboratory Medicine, University of Cincinnati College of Medicine, Ohio 45267-0529.
J Submicrosc Cytol Pathol. 1993 Apr;25(2):193-201.
Oxygen-derived free radicals are known to take part in cardiac injury during post-ischemic reperfusion (I/R). Xanthine oxidase (XO) is closely associated with the generation of superoxide radicals. We have determined the distribution of XO in rat myocardium after ischemia (I) and I/R by immunocytochemical method using murine monoclonal antibody against XO (bovine milk) and by enzyme histochemistry (EHC) in situ. Frozen sections of periodate-lysine-paraformaldehyde (PLP) fixed myocardium after 15, 60 and 90 min ischemia and 15 min ischemia and 30 min reperfusion were processed for immunocytochemistry and EHC. In other experiments, rats were treated with allopurinol, an inhibitor of XO, and hearts were processed for immunocytochemistry. By immunoperoxidase and immunofluorescence methods, a deep staining of interstitial cells, capillary and small blood vessels was observed, but the staining intensity of these cells was increased after reperfusion, in comparison to the normal and ischemic heart tissue. In the electron microscope, an immunoperoxidase reaction product was seen in the cytoplasm of interstitial, endothelial and smooth muscle cells. Similarly, EHC studies by nitroblue tetrazolium staining showed an increase in enzymatic activity in the tissue after reperfusion. The allopurinol-treated I/R tissue exhibited reduced staining. The data suggest that XO activity increases during ischemia but intensifies after reperfusion. The enzyme is localized in interstitial cells, coronary vessel endothelium and smooth muscle cells. XO is constantly present in the interstitial cells of the myocardium and it is a new finding not previously reported. It is further suggested that myocardial interstitium may be one of the major sites where oxygen derived radicals are generated during ischemia.
已知氧衍生的自由基在缺血后再灌注（I/R）期间参与心脏损伤。黄嘌呤氧化酶（XO）与超氧自由基的产生密切相关。我们通过使用抗XO（牛乳）的鼠单克隆抗体的免疫细胞化学方法和原位酶组织化学（EHC），确定了缺血（I）和I/R后大鼠心肌中XO的分布。对经高碘酸盐-赖氨酸-多聚甲醛（PLP）固定的心肌在缺血15、60和90分钟以及缺血15分钟和再灌注30分钟后的冰冻切片进行免疫细胞化学和EHC处理。在其他实验中，用XO抑制剂别嘌呤醇处理大鼠，并对心脏进行免疫细胞化学处理。通过免疫过氧化物酶和免疫荧光方法，观察到间质细胞、毛细血管和小血管有深染色，但与正常和缺血心脏组织相比，再灌注后这些细胞的染色强度增加。在电子显微镜下，在间质、内皮和平滑肌细胞的细胞质中可见免疫过氧化物酶反应产物。同样，通过硝基蓝四唑染色的EHC研究表明，再灌注后组织中的酶活性增加。经别嘌呤醇处理的I/R组织染色减少。数据表明，XO活性在缺血期间增加，但在再灌注后增强。该酶定位于间质细胞、冠状血管内皮和平滑肌细胞。XO持续存在于心肌的间质细胞中，这是一个以前未报道过的新发现。进一步表明，心肌间质可能是缺血期间产生氧衍生自由基的主要部位之一。
