Cubilin反义RNA对大鼠肾小管超负荷重吸收白蛋白的抑制作用

Inhibitory effect of Cubilin antisense RNA on albumin overload in renal tubules in rats

目的探讨经肾动脉灌注腺病毒载体介导Cubilin反义RNA对肾小管超负荷重吸收白蛋白的抑制作用。方法建立大鼠阿霉素肾病模型,正义RNA组和反义RNA组分别经肾动脉缓慢灌注GFP基因标记的pAdEasy-Cub和pAdEasy-ACub,对照组和模型组灌注等量生理盐水。激光共聚焦显微镜下观察GFP的表达,并监测24h尿蛋白量及其他生化指标;光镜下观察肾小管间质病理变化;免疫组化法观察肾小管Cubilin基因的表达以及肾小管上皮细胞内白蛋白的沉积。结果转染第4天,正、反义RNA组肾脏GFP基因有较强表达,11d达到高峰,可持续至实验结束;反义RNA组与其他各组相比,24h尿蛋白量明显增高(P<0.01),小管间质病变轻微,Cubilin阳性表达率最低,且与白蛋白的沉积呈显著正相关(r=0.896,P<0.01)。结论经肾动脉途径灌注重组腺病毒,能器官靶向性表达外源基因;Cubilin反义RNA对肾小管超负荷重吸收白蛋白有明显的抑制作用,并可以有效地减轻蛋白尿引起的肾间质损伤。

Objective To evaluate the inhibitory effect of Cubilin antisense RNA mediated by recombinant adenovirus vector on albumin overload in renal tubules in rats. Methods The rat models of adriamycin （ADR） induced nephritis were established. Cub vector group and ACub vector group underwent infusion via renal artery with pAdEasy-Cub, pAdEasy-ACub both marked by GFP gene respectively; control group and model group were given the same dose of normal saline in the same way. The GFP expression was observed by laser scanning confocal microscopy; the changes of uria protein in 24 hours and other biochemical indicators were measured simultaneously; the pathologic changes of renal tubule interstitium were observed under light microscope, and the Cubilin expression and the albumin deposition in renal tubules by immunohistochemistry. Results On the 4th day after transfection, the expression of GFP gene was detected in kidney in Cub vector group and ACub vector group, reached peak on the 11^th day, lasted to the end of experiment. Compared with other groups, urine protein in 24 hours in ACub vector group increased obviously （ P 〈 0. 01 ） , the pathologic changes of renal tubule interstitium were mildest, and the positive rate of Cubilin expression was the lowest. The significant positive correlation was found between Cubilin expression and albumin deposition （ r = 0. 896, P 〈0.01 ）. Conclusion When recombinant adenovirus vector was injected through renal artery, the introduced gene expressed preferentially in kidney. The transfected Cubilin antisense RNA has significant inhibitory effect on albumin overload in renal tubules and efficiently decreases the renal interstitial injury induced by proteinuria.