Medcalf J F, Walls J, Pawluczyk I Z, Harris K P
Department of Nephrology, Leicester General Hospital, Gwendolen Road, Leicester LE5 4PW, UK.
Nephrol Dial Transplant. 2001 Sep;16(9):1885-92. doi: 10.1093/ndt/16.9.1885.
Dialysate glucose has been implicated in the loss of peritoneal membrane function seen in long-term CAPD patients.
In order to investigate this in vitro, human peritoneal mesothelial cells (HPMC) were cultured in a 50:50 mix of dialysis solution and M199 for 12 h. The dialysate was laboratory manufactured and designed to be identical in composition to PD4 (LAB). The final glucose concentration ranged between 5 and 40 mmol/l. Experiments were conducted in the presence and absence of an anti-transforming growth factor-beta (TGF-beta) antibody. Cell viability was measured by lactate dehydrogenase (LDH) release. Fibronectin (FN) and TGF-beta protein were measured by ELISA, and FN gene expression was measured by Northern analysis. Separately, the effects of recombinant TGF-beta(1) added to M199: dialysate at 5 mmol/l glucose were investigated.
Forty millimoles per litre d-glucose LAB caused a decrease in cell viability, as evidenced by an increase in LDH release (6.0+/-1.3 vs 2.6+/-0.7%). This effect was dependent on osmolality. Forty millimoles per litre d-glucose LAB stimulated a 15.4+/-4.6% increase in FN, a 46.5+/-18.3% increase in TGF-beta protein (both P<0.05), and 1.4+/-0.09-fold increase in FN mRNA compared with 5 mmol/l d-glucose LAB. Exogenous TGF-beta 0-1 ng/ml induced a dose-dependent increase in FN protein (280+/-45% increase at TGF-beta 1 ng/ml, P<0.0001), and FN mRNA levels (10.0+/-1.8-fold at TGF-beta 1 ng/ml). The increase in FN in response to 40 mmol/l glucose was significantly reduced by anti-TGF-beta antibody to levels not different from control (93.8+/-6.6%, P<0.05 vs no Ab).
These data suggest that the pro-fibrotic effect of glucose dialysate on HPMC is mediated through stimulation of TGF-beta, which promotes FN gene expression and protein production.
透析液葡萄糖与长期持续性非卧床腹膜透析（CAPD）患者腹膜功能丧失有关。
为了在体外研究这一情况，将人腹膜间皮细胞（HPMC）在透析液与M199按50:50混合的培养液中培养12小时。透析液由实验室配制，其成分设计为与PD4（实验室产品）相同。最终葡萄糖浓度在5至40毫摩尔/升之间。实验在有和没有抗转化生长因子-β（TGF-β）抗体的情况下进行。通过乳酸脱氢酶（LDH）释放来测量细胞活力。通过酶联免疫吸附测定法（ELISA）测量纤连蛋白（FN）和TGF-β蛋白，通过Northern分析测量FN基因表达。另外，研究了向含5毫摩尔/升葡萄糖的M199:透析液中添加重组TGF-β(1)的效果。
40毫摩尔/升的d-葡萄糖实验室产品导致细胞活力下降，LDH释放增加证明了这一点（6.0±1.3%对2.6±0.7%）。这种作用取决于渗透压。与5毫摩尔/升的d-葡萄糖实验室产品相比，40毫摩尔/升的d-葡萄糖实验室产品刺激FN增加15.4±4.6%，TGF-β蛋白增加46.5±18.3%（两者P<0.05），FN mRNA增加1.4±0.09倍。外源性TGF-β 0至1纳克/毫升诱导FN蛋白呈剂量依赖性增加（在TGF-β 1纳克/毫升时增加280±45%，P<0.0001），以及FN mRNA水平增加（在TGF-β 1纳克/毫升时增加10.0±1.8倍）。抗TGF-β抗体使对40毫摩尔/升葡萄糖反应的FN增加显著降低至与对照无差异的水平（93.8±6.6%，与无抗体相比P<0.05）。
这些数据表明，葡萄糖透析液对HPMC的促纤维化作用是通过刺激TGF-β介导的，TGF-β促进FN基因表达和蛋白产生。
