Teng Yue-e, Hu Xiao-bin, Wei Ke-lun
Department of Medical Oncology, First Affiliated Hospital, China Medical University, Shenyang 110001, China.
Zhonghua Fu Chan Ke Za Zhi. 2003 Jun;38(6):337-9.
The aim of this study is to observe the kinetic changes of protein kinase C (PKC) activities in kidney of fetal rat after intrauterine distress (IUD), and to investigate the roles of PKC in the pathogenesis of kidney impair in IUD of fetal rat.
SD rats pregnant for 21 days were anesthetized intraabdominally, then one side of vessels of the two horns of uterus were occluded by arterial clamp to make models of intrauterine ischemia, hypoxia (IH) and reperfusion of fetal rat. The fetal rats were divided into IH group (ischemic and hypoxic for 15 min and 30 min, 16 fetal rats for each time point), reperfusion group (ischemic and hypoxic for 30 min, then reperfused for 15 min, 30 min, 45 min, 1 h and 2 h, respectively. 16 fetal rats for each time point) and false operation group (FOG) (18 fetal rats unclamped in the other uterus horn). The kidney tissue was homogenized first, then cell membrane protein and cytoplasmic protein were extracted. Improved Takai method was used to detect the changes of PKC activities of cell membrane and cytoplasma in kidney tissue of fetal rats.
(1) PKC activity of cell membrane started to increase after ischemia. The PKC activity of IH group was (5.66 +/- 0.88) pmol.mg(-1).min(-1); FOG was (4.30 + 0.97) pmol.mg(-1).min(-1). The PKC activity of IH group was significantly higher than that of FOG (P < 0.01). PKC activity of cell membrane increased rapidly after reperfusion, reached a peak at 30 min, and then decreased. The PKC activity was (7.26 +/- 0.76) pmol.mg(-1).min(-1) at 15 min, (9.25 +/- 0.94) pmol.mg(-1).min(-1) at 30 min, (8.34 +/- 0.89) pmol.mg(-1).min(-1) at 45 min, (6.57 +/- 0.96) pmol.mg(-1).min(-1) at 1 h, respectively. The PKC activity of reperfusion group was significantly higher than those of FOG (P < 0.01). The PKC activity of reperfusion group was (4.64 +/- 0.96) pmol.mg(-1).min(-1) at 2 h. There was no significant difference compared with that of FOG (P > 0.05). (2) The cytoplasmic PKC activity changed in a tendency opposite to that of the cell membrane PKC activity. In IH group, the PKC activity was (9.74 +/- 1.25) pmol.mg(-1).min(-1) at 15 min, (8.47 +/- 0.84) pmol.mg(-1).min(-1) at 30 min. FOG group was (10.63 +/- 1.92) pmol.mg(-1).min(-1). Reperfusion group reached the lowest PKC activity when reperfused for 30 min, (6.60 +/- 0.94) pmol.mg(-).min(-1), with a significant difference compared with that of FOG (P < 0.01). Then the PKC activity recovered to the level of FOG when reperfused for 2 h, (9.86 +/- 1.00) pmol.mg(-1).min(-1).
After intrauterine ischemia, hypoxia and reperfusion, PKC moves rapidly from cytoplasm to cell membrane in kidney tissue of fetal rat. PKC may be an important intracellular second messenger. It may play an important role in kidney impair in perinatal period.
本研究旨在观察宫内窘迫（IUD）后胎鼠肾脏中蛋白激酶C（PKC）活性的动态变化，探讨PKC在胎鼠IUD肾损伤发病机制中的作用。
将孕21天的SD大鼠腹腔麻醉，用动脉夹夹闭子宫两角一侧血管，制作胎鼠宫内缺血、缺氧（IH）及再灌注模型。将胎鼠分为IH组（缺血缺氧15分钟和30分钟，每个时间点16只胎鼠）、再灌注组（缺血缺氧30分钟，然后分别再灌注15分钟、30分钟、45分钟、1小时和2小时，每个时间点16只胎鼠）和假手术组（FOG）（另一子宫角未夹闭的18只胎鼠）。首先将肾组织匀浆，然后提取细胞膜蛋白和细胞质蛋白。采用改良的Takai法检测胎鼠肾组织细胞膜和细胞质中PKC活性的变化。
（1）缺血后细胞膜PKC活性开始升高。IH组PKC活性为（5.66±0.88）pmol·mg⁻¹·min⁻¹；FOG组为（4.30 + 0.97）pmol·mg⁻¹·min⁻¹。IH组PKC活性显著高于FOG组（P<0.01）。再灌注后细胞膜PKC活性迅速升高，30分钟时达到峰值，然后下降。15分钟时PKC活性为（7.26±0.76）pmol·mg⁻¹·min⁻¹，30分钟时为（9.25±0.94）pmol·mg⁻¹·min⁻¹，45分钟时为（8.34±0.89）pmol·mg⁻¹·min⁻¹，1小时时为（6.57±0.96）pmol·mg⁻¹·min⁻¹。再灌注组PKC活性显著高于FOG组（P<0.01）。再灌注2小时时，再灌注组PKC活性为（4.64±0.96）pmol·mg⁻¹·min⁻¹，与FOG组相比无显著差异（P>0.05）。（2）细胞质PKC活性变化趋势与细胞膜PKC活性相反。在IH组，15分钟时PKC活性为（9.74±1.25）pmol·mg⁻¹·min⁻¹，30分钟时为（8.47±0.84）pmol·mg⁻¹·min⁻¹。FOG组为（10.63±1.92）pmol·mg⁻¹·min⁻¹。再灌注组再灌注30分钟时PKC活性最低，为（6.60±0.94）pmol·mg⁻¹·min⁻¹，与FOG组相比有显著差异（P<0.01）。再灌注2小时时PKC活性恢复到FOG组水平，为（9.86±1.00）pmol·mg⁻¹·min⁻¹。
宫内缺血、缺氧及再灌注后，胎鼠肾组织中PKC迅速从细胞质转移至细胞膜。PKC可能是重要的细胞内第二信使，在围生期肾损伤中可能起重要作用。
