Indian Institute of Science Education and Research (IISER), Pune, Dr. Homi Bhabha Road, Pashan, Pune 411 008, India.
J Biosci. 2024;49.
Fibroblasts embedded in a 3D matrix microenvironment can remodel the matrix to regulate cell adhesion and function. Collagen hydrogels are a useful system to study cell-matrix interactions in a 3D microenvironment. While major matrix reorganizations are easily recognizable, subtle changes in response to environmental or biochemical cues are challenging to discern in 3D hydrogels. Three-dimensional collagen gels at 1.0 mg/ml vs 1.5 mg/ml were labelled with DQ-collagen and imaged by confocal reflectance microscopy to evaluate these small changes. An image analysis pipeline was developed, hydrogel area and number of crosssections analysed were optimized, and fibrillar collagen properties (number of branches, number of junctions, and average branch length) were quantified. While no significant changes were seen in fibrillar collagen organization between 1.0 mg/ml and 1.5 mg/ml collagen hydrogels, embedded mouse fibroblasts caused a significant increase in collagen branching and organization. Using the phalloidin-labelled cells, this change was quantitated in immediate proximity of the cell. A distinct increase in branch and junction numbers was observed, significantly altered by small changes in collagen concentration (1.0 mg/ml vs 1.5 mg/ml). Together, this analysis gives a quantitative evaluation of how cells respond to and modify their immediate microenvironment in a 3D collagen hydrogel.
嵌入在 3D 基质微环境中的成纤维细胞可以重塑基质，从而调节细胞黏附与功能。胶原水凝胶是研究 3D 微环境中细胞-基质相互作用的有用体系。虽然主要的基质重排很容易识别，但在 3D 水凝胶中，对于环境或生化线索的细微变化的识别则具有挑战性。以 1.0mg/ml 和 1.5mg/ml 的浓度标记 1.0mg/ml 和 1.5mg/ml 的胶原水凝胶，并通过共聚焦反射显微镜对其进行成像，以评估这些微小变化。开发了一种图像分析管道，优化了水凝胶面积和分析的横截面数量，并对纤维状胶原的特性（分支数量、连接数量和平均分支长度）进行了定量分析。尽管在 1.0mg/ml 和 1.5mg/ml 的胶原水凝胶之间，纤维状胶原组织没有明显变化，但嵌入的小鼠成纤维细胞导致胶原分支和组织显著增加。通过标记鬼笔环肽的细胞，在细胞的紧邻处定量了这种变化。观察到分支和连接数量明显增加，胶原浓度的微小变化（1.0mg/ml 与 1.5mg/ml）显著改变了这种变化。总之，这种分析为细胞如何响应并在 3D 胶原水凝胶中修饰其即时微环境提供了定量评估。
