缺血再灌注损伤后小鼠肾脏血管内皮生长因子C的表达

Expression of vascular endothelial growth factor C in the mice kidneys with ischemia reperfusion injury

目的探讨小鼠肾缺血再灌注损伤模型中血管内皮生长因子C的表达变化及其意义。方法建立小鼠肾缺血再灌注损伤模型。雄性C57BL／6小鼠用无创性动脉夹夹闭左肾动脉，置于32℃温箱后1h松开血管夹，取出右肾。Sham组操作同上，但不夹闭左肾动脉。再灌注0，6，12，24，48h后处死小鼠，收集外周血及肾脏标本。测定血肌酐（SCr）和尿素氮（BUN）水平。HE染色观察Sham组和缺血再灌注24h组（IR24h组）肾脏病理学变化，免疫组织化学检测Sham组和IR24h组血管内皮生长因子C（vascularendothelialgrowthfactorC，VEGF-C）在肾脏的表达及分布，连续切片检测VEGF-C与其受体血管内皮生长因子受体3（VEGFR-3）的共定位。Westernblot检测缺血后不同灌注时间VEGF-C的表达变化。结果小鼠肾脏缺血再灌注损伤后，SCr和BUN水平上升，且随着再灌注时间的延长肾功能损伤逐渐加重，再灌注24h时肾功能损伤最明显，再灌注48h时肾功能已经有部分恢复。与Sham组比较，缺血再灌注24h肾脏组织肾小管管腔扩张，内有管型形成，肾小管上皮细胞肿胀坏死，空泡变性，刷状缘坏死脱落，并且伴有炎性细胞侵润，而肾小球未见明显病变。免疫组织化学结果显示，与Sham组比较，缺血再灌注24h肾脏VEGF-C及其受体VEGFR-3的表达均明显增加，二者存在着共定位现象，且主要表达在肾脏皮髓质交界处及髓质部的肾小管。Westernblot结果显示随着缺血后再灌注时间的延长，VEGF-C的表达增加。结论肾缺血再灌注损伤后存在肾脏VEGF-C的表达上升，且与其受体VEGFR-3的表达增加呈共定位，推测VEGF-C可能参与了肾脏缺血再灌注损伤。

Objective To observe the changes of vascular endothelial growth factor C（VEGF-C） expression during renal ischemia/reperfusion and the clinical implication. Methods Renal ischemia reperfusion injury in male C57/BL6 mice was induced by clamping the left renal pedicles. The mice were remained in a 32 ℃ incubator for 1 h, and their right kidneys were removed. The mice in the sham group were given the same procedures except calmping the renal pedicles. These mice were sacrificed at 0,6,12,24 and 48 h after reperfusion and the kidneys and blood were collected. The levels of serum creatinine and blood urea nitrogen were determined. Renal pathological changes were evaluated by PAS staining. Immunohistochemistry was used to detect the distribution of VEGF-C and VEGFR3 and the colocalization of VEGF-C and VEGFR3 on serial sections. Western blotting was used to observe the dynamic expression changes of VEGF-C proteins at different reperfusion time points. Results The lev- els of serum creatinine and blood urea nitrogen were significatly increased in a time-dependent manner from 0 h to 48 h after reperfusion in mice. Notably, the renal function was partially restored at 48 h af- ter reperfusion. Renal pathological injury in IR24 h group was significantly aggravated as compared with the sham group. The percentage of renal tubular necrosis and proteincast was also significantly in- creased at 24 h after reperfusion. The expression of VEGF-C in IR24 h group was significantly higher than in the sham group, especially at the junction of renal cortex and medulla,and VEGF-C and VEG- FR3 were colocalizated in the renal tubules. The expression of VEGF-C was increased in a time-de- pendent manner confirmed by Western blotting. Conclusions VEGF-C and its receptor are up-regula- ted in the kidney tissues after renal ischemia/reperfusion, speculating that VEGF-C may play a role in the process of acute kidney injury.