冷保存对大鼠供肾结缔组织生长因子表达的影响

EFFECTS OF CRYOPRESERVATION ON EXPRESSION OF CONNECTIVE TISSUE GROWTH FACTOR IN RAT KIDNEY

目的探讨大鼠供肾在不同冷保存时间,其肾小管近端上皮细胞结缔组织生长因子(connective tissue growth factor,CTGF)的表达。方法健康SPF级Wistar雄性大鼠40只,体重230～250g。采取原位灌注切取整块双侧肾脏,置于0～4℃高渗枸橼酸盐嘌呤肾保存液中保存。根据保存时间不同,分为0、12、24、36、48h组(n=8)。采用免疫组织化学、原位杂交方法分别检测肾小管近端上皮细胞CTGF表达水平。结果在0、12h组大鼠供肾肾小管近端上皮细胞中CTGF极少表达,经图像分析两组CTGF蛋白阳性单位值(positive unit,PU)分别为5.91±2.30、5.57±2.40;CTGF mRNA PU值分别6.24±2.79、6.51±2.43,差异均无统计学意义(P>0.05)。随着冷缺血时间延长,24、36、48h组CTGF蛋白和CTGF mRNA表达均逐渐增加,24h组PU值分别为10.25±2.92、15.24±3.95;36h组分别为14.31±2.83、19.20±4.73;48h组分别为18.11±3.94、23.09±4.40;各组间比较差异均有统计学意义(P<0.05),与0、12h组比较差异亦均有统计学意义(P<0.05)。结论冷缺血损伤促进大鼠肾脏CTGF表达增加,可能在肾小管上皮细胞损伤再生与修复中起重要作用。

Objective To investigate the effect of cryopreservation （CP） on the expression of connective tissue growth factor （CTGF） in the renal tubular epithelial cells. Methods A total of 40 male Wistar rats （weighing 230-250 g） were used in this study. En bloc removal with in situ cooling both kidneys and hypertonic citrate adenine preservation solution were adopted. The rat kidney was be preserved 0, 12, 24, 36 and 48 hours at 0-4℃ （n=8）, respectively. The expression of CTGF of renal tubular epithelial cells was detected by using immunohistochemistry and in situ hybridization analysis. Results The expression of CTGF was less in CP 0 hour group and CP 12 hours group, the positive unit （PU） values of CTGF protein were 5.91 ± 2.30 and 5.57 ± 2.40 （P 〉 0.05）, respectively, and the PU values of CTGF mRNA were 6.24 ± 2.79 and 6.51 ± 2.43 （P 〉 0.05）, respectively. The PU values of CTGF protein increased at CP 24 hours group （10.25 ± 2.92）, CP 36 hours group （14.31 ± 2.83） and CP 48 hours group （18.11 ± 3.94, P 〈 0.05）, respectively, and the PU values of CTGF mRNA increased at CP 24 hours group （15.24 ± 3.95）, CP 36 hours group （19.20 ± 4.73） and CP 48 hours group （23.09 ± 4.40, P 〈 0.05）, respectively; showing significant differences when compared with CP 0 hour group and CP 12 hours group （P 〈 0.05）. Conclusion CTGF expression may increase with severe cold ischemia injury, and might play an important role in regeneration and repair of renal tubular epithelial cell injury.