Zoja C, Orisio S, Perico N, Benigni A, Morigi M, Benatti L, Rambaldi A, Remuzzi G
Mario Negri Institute for Pharmacological Research, Bergamo, Italy.
Lab Invest. 1991 Jan;64(1):16-20.
The present study was designed to assess whether human glomerular mesangial cells in culture express preproendothelin gene and whether endothelin gene expression in the mesangium is regulated by factors potentially released by inflammatory cells and platelets infiltrating the glomerular tuft during the course of various types of glomerulonephritis. For this purpose mesangial cells were incubated for 6 hours in the presence of absence of interleukin 1 beta (IL-1 beta), transforming growth factor-beta (TBF-beta), the thromboxane A2 analogue U-46619, and thrombin. Resting mesangial cells expressed a 2.3-kilobase mRNA on blot hybridization analysis with a human cDNA preproendothelin probe, indicating that this type of cells, in addition to glomerular endothelial cells, constitutively expresses endothelin gene. IL-1 beta did not change endothelin mRNA levels in respect to unstimulated mesangial cells. At variance, TGF-beta, U-46619, and thrombin had a marked effect on endothelin mRNA, stimulating a 3- to 8-fold increase over basal levels. Quantification of actin mRNA and analysis of the autoradiographic signals provided validation of the difference in the endothelin mRNA levels. Expression of preproendothelin mRNA in either resting or stimulated mesangial cells was associated with synthesis and release of the corresponding peptide in the cell supernatant as determined by a specific radioimmunoassay for endothelin. Endothelin production from IL-1 beta stimulated mesangial cells was not different from that of unstimulated cells, whereas a significant (p less than 0.01) increase in endothelin production was observed after cell stimulation with TGF-beta, U-46619, and thrombin. The demonstration that mesangial cells constitutively express mRNA for preproendothelin and release endothelin into culture medium, together with the finding that endothelin gene expression and production in mesangial cells are regulated by molecules potentially released at glomerular level during an inflammatory reaction may suggest that endothelin participates in the complex process of glomerular disease progression.
本研究旨在评估培养的人肾小球系膜细胞是否表达前内皮素原基因，以及在各种类型的肾小球肾炎病程中，肾小球系膜中内皮素基因的表达是否受浸润肾小球丛的炎性细胞和血小板可能释放的因子调控。为此，将系膜细胞在有或无白细胞介素1β（IL-1β）、转化生长因子-β（TGF-β）、血栓素A2类似物U-46619和凝血酶的情况下孵育6小时。用人类前内皮素原cDNA探针进行印迹杂交分析时，静止的系膜细胞表达一种2.3千碱基的mRNA，表明这类细胞除肾小球内皮细胞外，也组成性地表达内皮素基因。与未受刺激的系膜细胞相比，IL-1β未改变内皮素mRNA水平。与之不同的是，TGF-β、U-46619和凝血酶对内皮素mRNA有显著影响，使其水平比基础水平增加3至8倍。肌动蛋白mRNA的定量分析和放射自显影信号分析证实了内皮素mRNA水平的差异。通过内皮素特异性放射免疫测定法确定，静止或受刺激的系膜细胞中前内皮素原mRNA的表达与细胞上清液中相应肽的合成和释放相关。IL-1β刺激的系膜细胞产生的内皮素与未受刺激的细胞无差异，而在用TGF-β、U-46619和凝血酶刺激细胞后，观察到内皮素产生显著增加（p小于0.01）。系膜细胞组成性表达前内皮素原mRNA并将内皮素释放到培养基中，以及系膜细胞中内皮素基因表达和产生受炎症反应期间肾小球水平可能释放的分子调控这一发现，可能提示内皮素参与了肾小球疾病进展的复杂过程。
