姜黄素抑制脂多糖刺激的系膜细胞增殖及其白细胞介素1β和巨噬细胞趋化蛋白1基因的表达

Curcumin inhibited the proliferation and extracellular matrix production of human mesangial cells

目的 观察姜黄素对系膜细胞增殖以及对系膜细胞基质蛋白分泌的影响 ,并进一步探索姜黄素对系膜细胞白细胞介素 (IL 1β)和巨噬细胞趋化蛋白 (MCP 1)基因表达的改变。 方法 采用不同浓度姜黄素处理体外培养的人肾小球系膜细胞后 ,分别收集上清液及细胞 ,应用ELISA的方法测定上清液中胶原Ⅳ和Ⅲ的蛋白分泌量 ,MTT法测定系膜细胞活性 (吸光度值 ) ,RT PCR的方法测定系膜细胞IL 1β和MCP 1基因的相对表达量 ,以未加LPS及姜黄素和仅加LPS不加姜黄素作为对照组。结果 当姜黄素浓度大于或等于 6 2 5 μmol/L时 ,系膜细胞增殖明显受到抑制 (P <0 0 1) ,且表现为浓度依赖性。在基础状态下 ,系膜细胞分泌一定量的胶原Ⅳ和Ⅲ [(10± 9 13)ng/ml和 (2 9 5± 0 5 8)ng/ml],亦有MCP 1mRNA表达 [(42 1± 14 98) % ],但IL 1βmRNA几乎不表达 ;经LPS刺激后其胶原Ⅳ和Ⅲ分泌增加 [(138 75± 2 3 2 3)ng/ml和 (38 2 5± 5 38)ng/ml],同时IL 1β和MCP 1基因表达上调 [分别为 (16 91± 1 6 8) %和 (76 6± 6 5 9) % ],而姜黄素浓度为 4 μmol/L时即能明显抑制LPS刺激的系膜细胞胶原Ⅳ和Ⅲ的蛋白分泌 (P <0 0 5 ) ,且在高浓度时作用更为明显 ;同时姜黄素还能消除LPS上调系膜细胞IL 1β和MCP 1基因表达的作用 (P

Objective Glomerulosclerosis is characterized by extracellular matrix accumulative and is often associated with mesangial cell proliferation. Curcumin showed a protective effect on anti-glomerular basement membrane (anti-GBM) nephritis in vivo, although their cellular localization and mechanism of action is still unclear. In this study, a glomerular mesangial cell line derived from fetus was used to determine whether curcumin could inhibit the cell proliferation and alter the extracellular matrix turnover.Methods The cell activity was determined with MTT method. Mesangial cells were cultured in vitro and incubated with 0, 3.125, 6.25, 12.5, 25, 50, 100 and 200 μmol/L curcumin. In addition,human mesangial cells were cultured with or without LPS (10 μg/ml) in presence or absence of various concentrations of curcumin (4, 16 and 200 μmol/L), respectively. The supernatant and cells were collected. Then, the levels of the collagen type Ⅳ and Ⅲ protein in the supernatant were determined by using enzyme-linked immunosorbent assay and the IL-1β and MCP-1 mRNA in the cells was measured by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) after subconfluent quiescent mesangial cells were incubated with various concentrations of curcumin for 24 h in vitro.Results Curcumin at the concentration equal to or over 6.25 μmol/L was a ble to inhibit the proliferation of mesangial cells in a dose-dependent manner, the optical density according to the sequential concentrations of curcumin was 0.65±0.02, 0.62±0.04,0.56±0.01,0.53±0.02,0.51±0.03,0.44±0.05,0.41±0.07 and 0.38±0.06. Without any stimulation, human mesangial cells secreted some collagen type Ⅳ and Ⅲ (10±9.13 ng/ml and 29.5±0.58 ng/ml, respectively) and expressed some MCP-1 mRNA, but did not express IL-1β mRNA. LPS increased the expression of collagen typeⅣand Ⅲin the culture medium of mesangial cells in vitro [(138.75±23.23)ng/ml and (38.25±5.38) ng/ml ]and up-regulated the IL-1β and MCP-1 mRNA expression [ (16.91±1.68)% and (76.6±6.59)%]. Yet curcumin could significantly decrease collagen typeⅣandⅢin the supernatant of cultured mesangial cells induced by LPS (20.5±1.00, P<0.05 and 20.5±4.12 ng/ml, P<0.05) and down-regulated the mRNA expression of IL-1β and MCP-1 in mesangial cells induced by LPS (P<0.01).Conclusion Curcumin could inhibit the human mesangial cell proliferation and alter the extracellular matrix turnover, meanwhile it could down-regulate the IL-1β and MCP-1 mRNA expression induced by LPS, which may be valuable in decreasing the progression of glomerulosclerosis.