Astrab Leilani R, Skelton Mackenzie L, Caliari Steven R
Department of Biomedical Engineering, University of Virginia, Charlottesville, Virginia 22903.
Department of Chemical Engineering, University of Virginia, Charlottesville, Virginia 22903.
bioRxiv. 2024 Oct 15:2024.10.13.618034. doi: 10.1101/2024.10.13.618034.
Fibroblast activation drives fibrotic diseases such as pulmonary fibrosis. However, the complex interplay of how tissue mechanics and macrophage signals combine to influence fibroblast activation is not well understood. Here, we use hyaluronic acid hydrogels as a tunable cell culture system to mimic lung tissue stiffness and viscoelasticity. We applied this platform to investigate the influence of macrophage signaling on fibroblast activation. Fibroblasts cultured on stiff (50 kPa) hydrogels mimicking fibrotic tissue exhibit increased activation as measured by spreading as well as type I collagen and cadherin-11 expression compared to fibroblasts cultured on soft (1 kPa) viscoelastic hydrogels mimicking normal tissue. These trends were unchanged in fibroblasts cultured with macrophage-conditioned media. However, fibroblasts directly co-cultured with M2 macrophages show increased activation, even on soft viscoelastic hydrogels that normally suppress activation. Inhibition of interleukin 6 (IL6) signaling does not change activation in fibroblast-only cultures but ameliorates the pro-fibrotic effects of M2 macrophage co-culture. These results underscore the ability of direct M2 macrophage co-culture to override hydrogel viscoelasticity to promote fibroblast activation in an IL6-dependent manner. This work also highlights the utility of using hydrogels to deconstruct complex tissue microenvironments to better understand the interplay between microenvironmental mechanical and cellular cues.
成纤维细胞活化会引发诸如肺纤维化等纤维化疾病。然而，组织力学和巨噬细胞信号如何相互作用以影响成纤维细胞活化这一复杂过程尚未得到充分理解。在此，我们使用透明质酸水凝胶作为一种可调节的细胞培养系统来模拟肺组织的硬度和粘弹性。我们应用该平台来研究巨噬细胞信号对成纤维细胞活化的影响。与在模拟正常组织的柔软（1千帕）粘弹性水凝胶上培养的成纤维细胞相比，在模拟纤维化组织的坚硬（50千帕）水凝胶上培养的成纤维细胞，通过铺展以及I型胶原蛋白和钙黏蛋白-11表达来衡量，其活化程度有所增加。在用巨噬细胞条件培养基培养的成纤维细胞中，这些趋势没有变化。然而，即使在通常会抑制活化的柔软粘弹性水凝胶上，直接与M2巨噬细胞共培养的成纤维细胞也显示出活化增加。白细胞介素6（IL6）信号的抑制在仅成纤维细胞的培养中不会改变活化情况，但能改善M2巨噬细胞共培养的促纤维化作用。这些结果强调了直接的M2巨噬细胞共培养能够以IL6依赖的方式克服水凝胶的粘弹性，从而促进成纤维细胞活化。这项工作还突出了使用水凝胶来解构复杂组织微环境以更好地理解微环境机械和细胞信号之间相互作用的实用性。
