A2AR敲除对新生小鼠缺氧缺血脑区cyt C表达的影响

腺苷（adenosine）A2A受体（A2A receptor,A2AR）作为腺苷四种受体亚型之一,对新生鼠脑缺氧缺血的作用尚存在争议,探讨其作用机制将有助于新生儿缺氧缺血性脑病（hypoxic-ischemic encephalopathy,HIE）的临床治疗。为此,该实验观察了新生鼠脑缺氧缺血后A2AR敲除对其神经行为学的影响;采用TUNEL技术结合HE染色检测神经细胞凋亡;采用免疫组织化学法检测活化天冬氨酸特异性半胱氨酸蛋白酶3（caspase 3）及胞浆中细胞色素C（cytochrome C,cyt C）的表达。结果发现,A2AR敲除损伤了新生鼠神经行为功能,使神经细胞的凋亡和胞浆中cyt C的表达增加、caspase3活化增强。其中,在脑缺氧缺血后的1,3,7 d,神经细胞凋亡和caspase 3活化较野生型显著增加（P〈0.01）,而胞浆中cyt C的表达仅在脑缺氧缺血后的1,3 d显著增加（P〈0.01）,且其表达与神经细胞凋亡、caspase 3的活化均呈显著正相关。这提示,A2AR敲除后可能通过促使cyt C由线粒体释放出来增加新生鼠脑缺氧缺血后神经细胞的凋亡。

Effect of Adenosine A2A Receptor Knockout on the Expression of cyt C in Partial Cerebral Tissues After HIBD in Neonatal Mice

The role of adenosine A2A receptor （A2AR）, an important subtype of adenosine receptors, in neonatal mice after hypoxia-ischemia brain damage （HIBD） is still controversial. Studies about the mechanism of A2AR in HIBD will contribute to the clinical treatment of neonatal hypoxia-ischemia encephalopathy （HIE）. Our experiment observed the effect of A2AR knockout mice on neurological behavior. We used TUNEL assay combined with HE staining to detect neuronal apoptosis, also we used immunohistochemical assay to detect the expression of active caspase 3 and cytosolic cyt C after HIBD in neonatal mice. In our study, we found that A2AR knockout dam- aged neurobehavioral function of neonatal mice and increased the expression of apoptotic neuron, active caspase 3 and cytosolic cyt C. The differences of neuronal apoptosis between A2AR knockout mice and wild type mice were significant at 1, 3, 7 d after HIBD in neonatal mice, as well as the differences of active caspase 3 （P〈0.01）, while the significant differences of cytosolic cyt C between these two genetypes appeared at 1, 3 d after HIBD （P〈0.01）. The expressions of cytosolic cyt C and neuronal apoptosis were positively related, as well as the expressions of cytosolic cyt C and active caspase 3. This prompted that A2AR knockout possibly increased neuronal apoptosis by promoting cyt C released into the cytosolic after HIBD in neonatal mice.