Huang Wen-yan, Sun Hua, Pan Xiao-qin, Fei Li, Guo Mei, Bao Hua-ying, Chen Rong-hua, Jiang Xin-you
Department of Pediatrics, The Second Affiliated Hospital; The Center of Pediatric Nephrology, Nanjing Medical University, Nanjing 210011, China.
Zhonghua Er Ke Za Zhi. 2004 Jul;42(7):524-8.
Renal interstitial fibrosis is the final common pathway leading to end-stage renal failure for progressive renal disease of various types. The present study was undertaken to add to the knowledge on colchicine's antiinflammatory and antifibrotic properties confirmed by both human and experimental studies. As the main effector cells, fibroblasts have a central role in the pathogenesis of renal fibrosis. This study aimed to investigate the effects of colchicine on the synthesis and excretion of cytokines transforming growth factor-beta1 (TGF-beta1), interleukin-1beta (IL-1beta) and extracellular matrix (collagen III, collagen IV) in human renal fibroblast.
Various concentrations of colchicine (5.0 micromol/L, 10.0 micromol/L, 20.0 micromol/L, 40.0 micromol/L) were used to pretreat human embryo renal fibroblasts for 1 hour which were cultured in vitro and then stimulated by 10.0 microg/ml of lipopolysaccharide (LPS). After 18 hours, these fibroblasts and their supernatant were collected. The expression of TGF-beta1 mRNA, IL-1beta mRNA in the cells was studied by using RT-PCR, and the excretion of TGF-beta1, IL-1beta, collagen III and collagen IV by the fibroblasts was assessed by ELISA respectively.
(1) By pure stimulation with 10.0 micro g/ml LPS, the expression of TGF-beta1 mRNA and IL-1beta mRNA of fibroblasts was up-regulated 3 times (66.1 +/- 1.6 vs. 22.3 +/- 2.0, q = 590.5, P = 0.002) and 4.7 times (22.0 +/- 2.2 vs. 4.7 +/- 0.8, q = 106.8, P = 0.009), respectively. The protein excretion of TGF-beta1 and IL-1beta was remarkably increased as well compared with the control group [TGF-beta1: (516 +/- 14) pg/ml vs. (420 +/- 5) pg/ml (q = 80.3, P = 0.012), IL-1beta: (3.4 +/- 0.3) pg/ml vs. (0.3 +/- 0.1) pg/ml (q = 297.9, P = 0.003)]. (2) Colchicine could inhibit the expression of TGF-beta1 mRNA and protein in a dose-dependent manner. IL-1beta mRNA and protein were both up-regulated by colchicine. (3) LPS could not stimulate the excretion of extracellular matrix by fibroblasts, but the excretion of collagen III and collagen IV was down regulated by colchicine in a dose-dependent manner.
(1) The expression of TGF-beta1 mRNA and the excretion of TGF-beta1 protein in the fibroblasts was significantly suppressed by colchicine, while the expression of IL-1beta mRNA and the excretion of IL-1beta protein were enhanced. (2) Colchicine has significant inhibitory effect on the excretion of extracellular matrix such as collagen III and collagen IV in fibroblasts.
肾间质纤维化是各类进行性肾脏疾病导致终末期肾衰竭的最终共同途径。本研究旨在进一步了解秋水仙碱的抗炎和抗纤维化特性，这已得到人体和实验研究的证实。成纤维细胞作为主要效应细胞，在肾纤维化发病机制中起核心作用。本研究旨在探讨秋水仙碱对人肾成纤维细胞中细胞因子转化生长因子-β1（TGF-β1）、白细胞介素-1β（IL-1β）以及细胞外基质（III型胶原、IV型胶原）合成和分泌的影响。
采用不同浓度的秋水仙碱（5.0微摩尔/升、10.0微摩尔/升、20.0微摩尔/升、40.0微摩尔/升）对体外培养的人胚肾成纤维细胞进行预处理1小时，然后用10.0微克/毫升的脂多糖（LPS）刺激。18小时后，收集这些成纤维细胞及其上清液。采用逆转录聚合酶链反应（RT-PCR）研究细胞中TGF-β1 mRNA、IL-1β mRNA的表达，并用酶联免疫吸附测定（ELISA）分别评估成纤维细胞对TGF-β1、IL-1β、III型胶原和IV型胶原的分泌。
（1）单纯用10.0微克/毫升LPS刺激，成纤维细胞中TGF-β1 mRNA和IL-1β mRNA的表达分别上调3倍（66.1±1.6对22.3±2.0，q = 590.5，P = 0.002）和4.7倍（22.0±2.2对4.7±0.8，q = 106.8，P = 0.009）。与对照组相比，TGF-β1和IL-1β的蛋白分泌也显著增加[TGF-β1：（516±14）皮克/毫升对（420±5）皮克/毫升（q = 80.3，P = 0.012），IL-1β：（3.4±0.3）皮克/毫升对（0.3±0.1）皮克/毫升（q = 297.9，P = 0.003）]。（2）秋水仙碱能以剂量依赖方式抑制TGF-β1 mRNA和蛋白的表达。秋水仙碱使IL-1β mRNA和蛋白均上调。（3）LPS不能刺激成纤维细胞分泌细胞外基质，但秋水仙碱能以剂量依赖方式下调III型胶原和IV型胶原的分泌。
（1）秋水仙碱显著抑制成纤维细胞中TGF-β1 mRNA的表达和TGF-β1蛋白的分泌，同时增强IL-1β mRNA的表达和IL-1β蛋白的分泌。（2）秋水仙碱对成纤维细胞中III型胶原和IV型胶原等细胞外基质的分泌有显著抑制作用。
