Andratsch Manfred, Feifel Elisabeth, Taylor Lynn, O'Hayre Morgan, Schramek Herbert, Curthoys Norman P, Gstraunthaler Gerhard
Department of Physiology and Medical Physics, Innsbruck Medical University, Fritz-Pregl-Strasse 3, A-6020 Innsbruck, Austria.
Am J Physiol Renal Physiol. 2007 Sep;293(3):F846-53. doi: 10.1152/ajprenal.00139.2007. Epub 2007 Jun 27.
During systemic acidosis, renal proximal tubular cells exhibit enhanced rates of bicarbonate and ammonium ion synthesis and undergo extensive hypertrophy. The former adaptations are accomplished, in part, by increased expression of glutaminase (GA). LLC-PK(1)-FBPase+ cells, a gluconeogenic line of porcine kidney cells, exhibit a rapid activation of the ERK1/2 and p38 MAPK pathways and a two- to threefold increase in GA mRNA when transferred to acidic medium (pH 6.9). Transforming growth factor-beta (TGF-beta), a potent activator of MAPK and Smad signaling cascades, also causes extensive renal hypertrophy. Thus the potential role of TGF-beta in the renal response to metabolic acidosis was investigated. Western blot analyses established that in LLC-PK(1)-FBPase+ cells, TGF-beta activated the ERK1/2, p38 MAPK, and Smad1/5/8 pathways, but not the JNK and Smad2/3 pathways. Addition of TGF-beta to cells cultured in normal medium (pH 7.4) produced a steady increase in GA mRNA, resulting in a twofold induction after 18 h. Western blot analysis indicated that treatment with either TGF-beta or acidic medium resulted in an increased level of fibronectin. However, the effects of the two treatments on both GA mRNA and fibronectin levels occurred with different time courses and were additive. In addition, the rates of ammonia production were decreased slightly by addition of TGF-beta. Finally, a GA-luciferase reporter construct, which is activated 3.5-fold by treatment with acidic medium, is not affected by TGF-beta. Therefore, TGF-beta and metabolic acidosis activate some of the same signaling pathways in LLC-PK(1)-FBPase+ cells, but produce separate effects on GA expression.
在全身性酸中毒期间，肾近端小管细胞表现出碳酸氢盐和铵离子合成速率增强，并发生广泛的肥大。前一种适应性变化部分是通过谷氨酰胺酶（GA）表达增加来实现的。LLC-PK(1)-FBPase+细胞是猪肾细胞的糖异生细胞系，当转移至酸性培养基（pH 6.9）时，表现出ERK1/2和p38丝裂原活化蛋白激酶（MAPK）途径的快速激活以及GA mRNA增加两到三倍。转化生长因子-β（TGF-β）是MAPK和Smad信号级联的有效激活剂，也会导致广泛的肾肥大。因此，研究了TGF-β在肾脏对代谢性酸中毒反应中的潜在作用。蛋白质印迹分析表明，在LLC-PK(1)-FBPase+细胞中，TGF-β激活了ERK1/2、p38 MAPK和Smad1/5/8途径，但未激活JNK和Smad2/3途径。向在正常培养基（pH 7.4）中培养的细胞中添加TGF-β会使GA mRNA稳步增加，18小时后诱导增加两倍。蛋白质印迹分析表明，用TGF-β或酸性培养基处理均导致纤连蛋白水平升高。然而，两种处理对GA mRNA和纤连蛋白水平的影响具有不同的时间进程且具有累加性。此外，添加TGF-β会使氨生成速率略有降低。最后，用酸性培养基处理可激活3.5倍的GA-荧光素酶报告基因构建体不受TGF-β影响。因此，TGF-β和代谢性酸中毒在LLC-PK(1)-FBPase+细胞中激活了一些相同的信号通路，但对GA表达产生了不同的影响。
