| dc.contributor.advisor | Lutz, Susanne Prof. Dr. | |
| dc.contributor.author | Santos, Gabriela Leão | |
| dc.date.accessioned | 2021-08-09T11:35:20Z | |
| dc.date.available | 2021-10-24T00:50:08Z | |
| dc.date.issued | 2021-08-09 | |
| dc.identifier.uri | http://hdl.handle.net/21.11130/00-1735-0000-0008-58D5-5 | |
| dc.identifier.uri | http://dx.doi.org/10.53846/goediss-8721 | |
| dc.language.iso | eng | de |
| dc.rights.uri | http://creativecommons.org/licenses/by-nc-nd/4.0/ | |
| dc.subject.ddc | 610 | |
| dc.title | Mechanical regulation of cardiac fibroblasts. | de |
| dc.type | doctoralThesis | de |
| dc.contributor.referee | Schu, Peter Prof. Dr. | |
| dc.date.examination | 2020-10-26 | |
| dc.description.abstracteng | Cardiac fibrosis progressively contributes to heart failure (HF). Yet, its complex nature still lacks a more comprehensive understanding. One main hurdle is the lack of representative human models providing insights into the behaviour of cardiac fibroblasts (CF) and their pathological derivatives. Therefore, our group developed a human engineered connective tissue (hECT) model based on primary human CF and collagen type 1, which allows studying the pathological CF conversion. In this model, the environmental stiffness provided by the mould’s architecture was used as the single trigger for the CF conversion. The moulds contained either two flexible poles (flexible model) or a single central stiff rod (stiff model). Destructive tensile strength measurements showed that flexible hECT model achieved a stiffness similar to that of the healthy myocardium, while stiff hECT model achieved a stiffness comparable to the diseased myocardium. In addition, stiff hECT showed lower elasticity and extensibility compared to flexible hECT, but could withstand higher stress. A gradual increase in the stiffness of the flexible poles further demonstrated to result in an incremental increase in tissue stiffness and a decrease in tissue contraction. Furthermore, the modulation of the internal cell stiffness by interfering with the actin-myosin cytoskeleton was investigated. To that purpose, ROCK inhibitors and Latrunculin A were applied to both hECT and rat engineered connective tissue (rECT) generated with neonatal rat cardiac fibroblasts. The data indicates that the integrity of the actin-myosin network is relevant for compaction, contraction and stiffening of ECT. The underlying pathway involved the MRTF/SRF-dependent regulation of gene expression, e.g., of the collagen cross-linker enzyme lysyl oxidase (LOX).
To obtain a more profound insight into the stiffness-dependent cellular remodelling, a comparison was conducted between 2D-cultured myofibroblasts and cells from flexible and stiff hECT. It was shown that a partial reversion of the myofibroblasts phenotype of 2D-cultured cells is achievable in hECT, as demonstrated by cell cycle activity and expression of myofibroblast markers. Importantly, the direct comparison of cells in flexible and stiff hECT showed a clear time-dependent segregation in two different cellular phenotypes. By RNA sequencing it was possible to demonstrate that the gene program differs between cells embedded in flexible and in stiff hECT, especially in gene clusters assigned to extracellular matrix organization and protein folding. In comparison with published data from human samples, hECT exposed to high environmental stiffness shows a transcriptome profile similar to that of a failing human heart.
In summary, this novel engineered connective tissue model recapitulates the mechanical changes that occur during fibrogenesis and assisted phenotypic switching of cardiac myo- and fibroblasts mediated by environmental stiffness. Thus, the hECT model opens new perspectives and offers a versatile alternative to study mechanisms underlying scarring of the human heart and for screening for anti-fibrotic drugs. | de |
| dc.contributor.coReferee | Dressel, Ralf Prof. Dr. | |
| dc.contributor.thirdReferee | Meyer, Thomas Prof. Dr. | |
| dc.contributor.thirdReferee | Katschinski, Dörthe Prof. Dr. | |
| dc.contributor.thirdReferee | Zeisberg, Michael Prof. Dr. | |
| dc.subject.eng | cardiac fibrosis | de |
| dc.subject.eng | cardiac fibroblasts | de |
| dc.subject.eng | myofibroblasts transdifferentiation | de |
| dc.subject.eng | mechanotransduction | de |
| dc.subject.eng | tissue engineering | de |
| dc.subject.eng | biomechanical tissue properties | de |
| dc.subject.eng | drug screening | de |
| dc.identifier.urn | urn:nbn:de:gbv:7-21.11130/00-1735-0000-0008-58D5-5-8 | |
| dc.affiliation.institute | Medizinische Fakultät | |
| dc.subject.gokfull | Medizin (PPN619874732) | de |
| dc.subject.gokfull | Pharmakologie (PPN619875518) | de |
| dc.subject.gokfull | Molekularbiologie {Medizin} (PPN619875186) | de |
| dc.description.embargoed | 2021-10-24 | |
| dc.identifier.ppn | 1765981875 | |