Damage to our tissues occurs daily and must be repaired by the body in a timely manner in order to prevent infection and restore tissue integrity. Many cell types are involved in the healing process, but it is the cells of the stroma that are largely responsible for rebuilding fibrous tissue, which provides structure and support for all other cell types during healing. This dissertation focuses on stromal tissue repair, the rebuilding of fibrous tissue by fibroblasts following injury. Specifically, I focus on 1) models to study wound healing in vitro, and the specific biological processes of healing that each model captures, 2) the response of engineered stromal microtissues to different methods of injury, namely laceration and laser ablation, and the subsequent clearance and rebuilding of the extracellular matrix by fibroblasts, and 3) how different types of stromal cells and extracellular matrix proteins contribute to tissue repair in vitro.

Degenerated intervertebral discs (IVDs) were treated with autologous adipose-derived stem cells (ASC) loaded into an injectable collagen scaffold in a sheep model to investigate the implant's therapeutic potential regarding the progression of degeneration of previously damaged discs. In this study, 18 merino sheep were subjected to a 3-step minimally invasive injury and treatment model, which consisted of surgically induced disc degeneration, treatment of IVDs with an ASC-loaded collagen hydrogel 6 weeks post-operatively, and assessment of the implant's influence on degenerative tissue changes after 6 and 12 months of grazing. Autologous ASCs were extracted from subcutaneous adipose tissue and cultivated in vitro. At the end of the experiment, disc heights were determined by µ-CT measurements and morphological tissue changes were histologically examined.Histological investigations show that, after treatment with the ASC-loaded collagen hydrogel implant, degeneration-specific features were observed less frequently. Quantitative studies of the degree of degeneration did not demonstrate a significant influence on potential tissue regeneration with treatment. Regarding disc height analysis, at both 6 and 12 months after treatment with the ASC-loaded collagen hydrogel implant a stabilization of the disc height can be seen. A complete restoration of the intervertebral disc heights however could not be achieved.The reported injection procedure describes in a preclinical model a translational therapeutic approach for degenerative disc diseases based on adipose-derived stem cells in a collagen hydrogel scaffold. Further investigations are planned with the use of a different injectable scaffold material using the same test model.

Under senaste decennierna, har intresset kring implantat från naturliga och syntetiska polymerer ökat markant i samband med en ökad marknadsefterfrågan på vävnadsdonationer. Detta har lett till efterforskningen av nya in-situ formerande geler med formbevarande egenskaper in-vivo. Extracellulära matrisen (ECM) innehåller flertal makromolekylära komponenter med stödjande och nätverksformerande egenskaper, då de ofta är essentiella strukturella konstituenter i biologiska system. Den huvudsakliga beståndsdelen i ECM-nätverket, kollagen typ-1, har undersökts som en kandidat för utvecklingen av nya modifierade biomaterial med cellförökande-, biokompatibla-, icke-svällande samt injicerbara egenskaper. I detta projekt var grisderiverat kollagen modifierat med furfuryl glycidyl eter, och tvärbundet med 10 kDa 8-armad PEG-malimid, vilket genomgick kovalenta Diels-alder klick-reaktioner. Fyra formuleringar användes i experimenten, baserat på de stökiometriska förhållandet mellan furan och malimid i det kemiskt modifierade kollagenet. Dessa kollagen-baserade hydrogeler undersöktes baserat på 4 wt% löst kollagen, med avseende på reologiska-, mekaniska-, bionedbrytbara och svällningsegenskaper.  Resultaten indikerar att en ökning i fastfas-mängd ledde till en förhöjning i hydrogelens styvhet. Detta kunde observeras genom en ökad lagringsmodul (G’) under reologiska mätningar. Samtidigt indikerade mätningarna att sprödheten av hydrogelen ökade i korrelation med ökningen av styvheten.  Vidare drogs slutsatsen att kovalenta interaktionerna är enbart delvis ansvarig för ökningen av G’. Jämförelser med tan delta och kritiska töjningen visade att det fanns fysiska interaktioner mellan polymererna vilket också bidrog till ökningen av G’ för gelformuleringar som innehöll furan-till-malimid förhållanden på 1:1 och 1:4. Dessa fysiska interaktioner tros härstamma från en ökning av hydrofobiska effekter mellan kollagen kedjorna, då agglomerering och löslighetssvårigheter i vattenlösningar observerades i flertal experiment. Kollagen-PEG-Malimid hydrogelen var också injicerbar genom 15G kanyler, nedbrytbar in-vitro i närvaro av kollagenas, och uppvisade låg svällning i vatten. Inga cellexperiment genomfördes, och därav kunde inga slutsatser dras i hydrogelens cellförökande egenskaper. Däremot har tidigare arbete av Dr Jamadi visat att kollagen-PEG-malimid hydrogel med 2 vikt% haft kapacitet att inkapsla celler. Detta kan vara en indikation att högre viktprocent av gelen också kan uppvisa samma effekt vid framtida försök.Sammanfattningsvis, kunde slutsatsen dras att hydrogelen uppvisar flertal av de spekulerade, samt några av de eftertraktade egenskaperna hos en injicerbar hydrogel som potentiellt kan användas kliniskt. / In recent decades, the interest in implants manufactured from natural and synthetic polymers has grown as the demand for tissue donations has increased. This process has led to the pursuit of new, in-situ forming gels with shape-retaining properties in-vivo. The extracellular matrix (ECM) contains several macromolecular constituents with scaffold forming capabilities and is an inherent part of the body. The main component in the ECM-scaffold, collagen type-I, has been investigated as a candidate for novel modified biomaterials with cell proliferating, biocompatible, non-swelling, and injectable properties. Collagen was modified with furfuryl glycidyl ether and crosslinked with 10 kDa 8-arm PEG-maleimide, which undergoes Diels-alder covalent click-type reactions. Four formulations were used, based on a stoichiometric ratio of furan to maleimide (1:1-1:4). These materials' properties were evaluated at 4 wt% collagen for rheological-, mechanical-, biodegradability and swelling characteristics. The results indicated that an increase in solid content improved stiffness in the hydrogel.  This was observed by an increase of storage modulus (G’) during rheological measurements. The same measurements also indicated that the hydrogel showed an increase in brittle characteristics correlated with higher solid content.  Furthermore, it was concluded that the covalent interactions are partly responsible for the increase of G’. Comparisons in tan delta and critical strain showed that there are physical interactions that cause the increase in moduli for gel formulations containing furan to maleimide ratios of 1:1 and 1:4. These physical interactions are thought to stem from the increase in hydrophobic effects of the modified collagen, as agglomeration and solubility issues in aqueous solutions are observed in multiple experiments.  Collagen-PEG-maleimide hydrogel was also injectable through a 15-gauge needle, degradable in-vitro, and showed low swelling. No cell experiments were performed, and hence no conclusions could be made of this aspect of the hydrogel. However, work has been performed by Dr Jamadi, which indicates that the Collagen-PEG-Maleimide hydrogel with lower weight percentages allows for cell encapsulation. Hence, it could be concluded that several characteristics of tissue mimetic material were met with this hydrogel.