Modulation of Osteogenic Cell Morphology by ECM Ligands and Enamel Matrix Derivative

Preduvjet za uspješnu regeneraciju parodontnog tkiva jest odgovarajuća aktivacija populacije za to odgovornih stanica, poput osteoblasta. U tom su slučaju stanična adhezija i sazrijevanje usko povezani s morfologijom stanica i f-aktinskom organizacijom citoskeleta. Mogućnost pojačavanja parodontalnog cijeljenja pomoću pojedinih komponenti izvanstaničnog matriksa (ECM) i derivata caklinskog matriksa dobro je dokumentirana u literaturi. Svrha rada: Svrha ovog istraživanja bila je testirati učinak ECM-bjelančevina, kolagena tipa I, laminina-1 te komercijalnog proizvoda EMD-a na morfološku i citoskeletnu organizaciju osteogenih stanica. Ispitanici i postupci: Tijekom promatranja analizirano je ukupno 2450 osteogenih stanica iz pet različitih staničnih linija (četiri primarnih i jedne komercijalne) kultiviranih na pojedinim supstratima, a analizirali su ih tri neovisna promatrača. Nakon bojenja za f-aktin staničnog citoskeleta i automatizirane CLSM-vizualizacije, stanice su podijeljene u tri skupine ovisno o njihovim morfološkim svojstvima (nezrele, prijelazne i zrele). Osim deskriptivne analize obavljena je bila i multivarijantna logička regresija radi identificiranja odgovarajućih parametara koji utječu na staničnu morfologiju i organizaciju citoskeleta. Rabljeni pojedinačni ligandi kolagena i laminina te posebice EMD poticali su stvaranje zrelog osteogenog fenotipa, premda su bile uočene i određene razlike među korištenim staničnim linijama. Rezultati: Analiza morfologije i citoskeleta pouzdan je način skupljanja prvih podataka o biokompatibilnosti i bioaktivaciji stanica na različitim supstratima. Naši rezultati upućuju na mogući potencijal istraživanih liganada u pojačavanju osteogenoga staničnog pričvrstka i sazrijevanju te time i pomaganju cijeljenja parodontnog tkiva.The precondition for successful periodontal regeneration is adequate activation of relevant cell populations like osteogenic cells. Here, cell adhesion and maturation are closely associated with cell morphology and f-actin cytoskeletal organisation. The potential of solitaire extracellular matrix (ECM) components as well as enamel matrix derivative (EMD) to enhance periodontal healing is well documented. Objective: The aim of the study was to test the impact of the ECM proteins collagen type 1 and laminin-1 as well as commercially available EMD on osteogenic cell morphology and cytoskeletal organisation. Material and methods: In an observational study, a total of 2450 osteogenic cells of 5 different cell lines (4 primary ones and 1 commercial one) cultivated on the respective substrates were analysed by 3 independent observers. After staining for the f-actin cytoskeleton and automated CLSM visualisation, cells were assigned to 3 different categories depending on morphological cell attributes (immature vs. intermediate vs. mature). Besides descriptive analysis, a multivariate logistic regression was performed to identify relevant influence parameters on cell morphology and cytoskeletal organisation. Results: The applied solitaire ligands collagen and laminin and especially EMD promoted a mature osteogenic phenotype. Nevertheless, considerable differences between the investigated cell lines could be identified as well. Analysis of cell morphology and cytoskeletal organisation offers a reliable method of acquiring the first hints of biocompatibility and bio-activation on different substrates. Conclusion: Our results highlight the potential of the investigated ligands to support periodontal regeneration by enhancing osteogenic cell attachment and maturation