Use of a three-dimensional cell culture model to study airway smooth muscle-mast cell interactions in airway remodeling
Use of a three-dimensional cell culture model to study airway smooth muscle-mast cell interactions in airway remodeling
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DOI：
10.1152/ajplung.90445.2008
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发表时间：
2009-06-01
影响因子：
4.9
通讯作者：
Johnson, Simon R.
中科院分区：
文献类型：
--
作者：
Ceresa, Claudia C.;Knox, Alan J.;Johnson, Simon R.
Ceresa CC，Knox AJ，约翰逊SR。使用三维细胞培养模型研究气道重塑中气道平滑肌-肥大细胞相互作用。Am J Physiol Lung Cell Mol Physiol 296：L1059-L1066，2009.首次发表于2009年4月3日; doi：10.1152/ajplung.90445.2008。气道平滑肌（ASM）质量增加和肥大细胞浸润是哮喘气道重塑的关键特征。我们描述了一个模型来研究ASM，细胞外基质，肥大细胞和气道重塑之间的关系。将ASM细胞单独或与肥大细胞一起在三维（3-D）胶原I凝胶（3-D培养物）中培养。免疫细胞化学和蛋白质印迹法的ASM在3-D文化揭示了一个纺锤形的形态和显着较低的α-平滑肌肌动蛋白和波形蛋白的表达比ASM培养在单层胶原蛋白I型或塑料（2-D文化）。在3-D培养中，通过Ki 67免疫细胞化学检测的基础ASM增殖降低至2-D培养中的33 +/- 7%（P < 0.05）。肥大细胞的存在使ASM的增殖增加了1.8倍（P < 0.05）。明胶酶谱显示更多的活性基质金属蛋白酶（MMP）-2在3-D比2-D培养上清液超过7天。通过凝胶收缩检查功能性MMP活性。MMP抑制剂伊洛马司他显著抑制了7天以上的自发凝胶收缩。肥大细胞共培养增强ASM凝胶收缩22 +/- 16%（不显著）。我们的模型显示，ASM具有不同的形态，在三维培养中具有较低的收缩蛋白表达和基础增殖。与标准技术相比，ASM合成功能，如MMP的产生和活性所示，持续更长的时间。肥大细胞的存在下，在3-D模型增强ASM增殖和MMP的生产。我们的系统可能比标准培养系统更准确地模拟哮喘中的气道重塑。
Ceresa CC, Knox AJ, Johnson SR. Use of a three-dimensional cell culture model to study airway smooth muscle-mast cell interactions in airway remodeling. Am J Physiol Lung Cell Mol Physiol 296: L1059-L1066, 2009. First published April 3, 2009; doi: 10.1152/ajplung.90445.2008.-Increased airway smooth muscle (ASM) mass and infiltration by mast cells are key features of airway remodeling in asthma. We describe a model to investigate the relationship between ASM, the extracellular matrix, mast cells, and airway remodeling. ASM cells were cultured in a three-dimensional (3-D) collagen I gel (3-D culture) alone or with mast cells. Immunocytochemistry and Western blotting of ASM in 3-D cultures revealed a spindle-shaped morphology and significantly lower alpha-smooth muscle actin and vimentin expression than in ASM cultured in monolayers on collagen type I or plastic (2-D culture). In 3-D cultures, basal ASM proliferation, examined by Ki67 immunocytochemistry, was reduced to 33 +/- 7% (P < 0.05) of that in 2-D cultures. The presence of mast cells in cocultures increased ASM proliferation by 1.8-fold (P < 0.05). Gelatin zymography revealed more active matrix metalloproteinase (MMP)-2 in 3-D than in 2-D culture supernatants over 7 days. Functional MMP activity was examined by gel contraction. The spontaneous gel contraction over 7 days was significantly inhibited by the MMP inhibitor ilomastat. Mast cell coculture enhanced ASM gel contraction by 22 +/- 16% (not significant). Our model shows that ASM has different morphology, with lower contractile protein expression and basal proliferation in 3-D culture. Compared with standard techniques, ASM synthetic function, as shown by MMP production and activity, is sustained over longer periods. The presence of mast cells in the 3-D model enhanced ASM proliferation and MMP production. Airway remodeling in asthma may be more accurately modeled by our system than by standard culture systems.