The Effect of Stimulation with Vascular Endothelial Growth Factor on Phenotype and Function of Adipose-derived Stromal Cells from Patients with Ischemic Heart Disease
Author
Larsen, Bjarke Follin
Term
4. term
Publication year
2012
Submitted on
2012-06-11
Pages
45
Abstract
Stamceller fra fedtvæv (ADSC) undersøges som behandling af iskæmisk hjertesygdom, hvor nedsat blodforsyning skader hjertet. Før injektion forbehandles cellerne ofte med vaskulær endothelvækstfaktor (VEGF), et signalstof der fremmer dannelse af blodkar, for at skubbe dem i retning af endothelceller (de celler, der beklæder blodkar). Dette studie undersøgte, hvordan VEGF påvirker ADSC fra hjertesyge: om behandlingen skubber dem mod en endothel-identitet, og om de bevarer deres mesenkymale stamcellettræk. Vi dyrkede ADSC med VEGF-A165 i 1, 2 eller 3 uger. Kontroller blev holdt enten i serumfattigt medium (få vækstfaktorer) eller i fuldt medium. Vi målte genaktivitet og protein-niveauer af VEGF-receptorer samt markører for endothel- og stamceller med standardmetoder (RT-qPCR, immunocytokemi og flowcytometri). Vi testede også, om cellerne kunne danne karlignende rør på en gel (ECMatrix). ADSC dyrket med VEGF under serumfattige forhold havde højere genudtryk af FOXF1 og PDGFβ sammenlignet med celler i fuldt medium. Andre endothel-gener (vWF, VEGFR1, VEGFR2) viste stigende tendenser, men uden statistisk signifikans. Endothelmarkører sås kun sporadisk ved immunfarvning og blev ikke bekræftet med flowcytometri. Rørdannelse in vitro sås kun med VEGF under serumfattige forhold. Mesenkymale stamcellemarkører var uændrede på tværs af betingelser. Samlet set syntes serumfattig kultur at forberede ADSC mod en endothel-retning, og VEGF gav ikke en tydelig ekstra effekt. Ændringerne var beskedne, idet kun den tidligste endothelmarkør var signifikant på genniveau. Serumsult gjorde cellerne mere tilbøjelige til at danne rør og ændrede ikke profilen af mesenkymale markører. VEGF kan medvirke til at forberede ADSC til endothel-differentiering, men yderligere signaler er nødvendige for at fuldende processen.
Fat-derived stem cells (ADSCs) are being explored as a therapy for ischemic heart disease, where poor blood supply damages the heart. Before injection, the cells are often exposed to vascular endothelial growth factor (VEGF), a signal that promotes blood vessel growth, to nudge them toward becoming endothelial cells (the cells that line blood vessels). This study examined how VEGF treatment affects ADSCs from heart patients: whether it pushes them toward an endothelial identity and whether they keep their mesenchymal stem cell features. We grew ADSCs with VEGF-A165 for 1, 2, or 3 weeks. Controls were kept either in serum-deprived medium (low growth factors) or in complete medium. We measured gene activity and protein levels of VEGF receptors and endothelial and stem cell markers using standard methods (RT-qPCR, immunocytochemistry, and flow cytometry). We also tested whether the cells could form vessel-like tubes on a gel (ECMatrix). ADSCs grown with VEGF under serum-deprived conditions showed higher gene expression of FOXF1 and PDGFβ compared with cells in complete medium. Other endothelial genes (vWF, VEGFR1, VEGFR2) trended upward but did not reach significance. Endothelial markers were seen only sporadically by immunostaining and were not confirmed by flow cytometry. Tube formation in vitro occurred only with VEGF under serum deprivation. Mesenchymal stem cell markers were unchanged across conditions. Overall, serum-deprived culture primed ADSCs toward an endothelial direction, and VEGF did not add a clear extra effect. The changes were subtle, with only the earliest endothelial marker reaching significance at the gene level. Serum deprivation made cells prone to form tubes and did not alter mesenchymal marker profiles. VEGF may help predispose ADSCs toward endothelial differentiation, but additional cues are needed to complete the process.
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