297 General discussion and future perspectives models. The differences between the ECM components with respect to content and amount in different regions of lung ECM in health and disease were described in a recent study using mass spectrometry by Hoffman et al [12]. An intriguing question that remains is to what extent lung ECM-derived hydrogels can recapitulate this protein content and composition. Analyzing lung ECM-derived hydrogels using similar approaches can strengthen arguments regarding the capacity of lung ECM-derived hydrogels to recapitulate the biochemical composition of lung ECM. Moreover, comparing the changes in proteomic profiles of the stroma through such analysis can generate novel findings. Another question that requires future research is whether cell-seeded lung ECM-derived hydrogels can re-capture, and expand upon, previous observations with respect to the applicability of ECM fragments as biomarkers of fibrotic lung diseases. Furthermore, approach to identify novel mechanisms of cell:ECM interactions in fibrosis using in vitro models developed in this thesis, could be through transcriptomics analysis. Profiling of healthy and IPF lungs using single-cell RNAsequencing (scRNAseq) has been extensively demonstrated in the last decade [18, 20, 22, 43-48]. By investigating the gene profiles of cells seeded in control or diseased lung ECM-derived hydrogels and comparing these gene signatures to the existing fibrotic RNA signatures, the potential applicability of the model to reflect gene changes that take place in the lung tissue could be revealed. After this initial validation, the utilization of the model can be expanded for assessing possible treatment preclinically. However, isolating RNA from cells, especially fibroblasts, from (human) lung ECM-derived hydrogels remains challenging due to poor quality and/or quantity of isolated RNA that is achievable to date. Although recovering cells via enzymatic digestion is an alternative, the cells might change their transcriptomic profile during the recovery process. Together with advances in the field of spatial transcriptomics, these challenges will need to be bypassed for a thorough genelevel characterization of cells embedded in lung ECM (diseased) microenvironment. Advanced imaging methodologies can also be applied to the in vitro models described in this thesis in order to promote our understanding on fibrotic ECM. Although the current field of clinical imaging is still limited to investigate the changes in ECM during lung fibrosis, novel methods such as optical coherence tomography (OCT), confocal laser endomicroscopy (CLE) or high resolution computed tomography (HRCT) to investigate ECM structure in vivo are being developed thanks to the advances in the fields of physics and imaging [49]. Some of the other methods such as photoacoustic or ultrasound imaging can also provide additional information about the tissue architecture [50]. Most of the deep tissue imaging methods so 10
RkJQdWJsaXNoZXIy MTk4NDMw