Angiogenesis, the formation of new blood vessels from existing ones, is an essential process for successful bone regeneration. Further, angiogenesis is a key factor for the development of bone-related disorders like osteosarcoma or arthritis. Fucoidans, sulfated polysaccharides from brown algae, have been shown to affect angiogenesis as well as a series of other physiological processes including inflammation or infection. However, the chemical properties of fucoidan which define the biological activity vary tremendously, making a prediction of the bioactivity or the corresponding therapeutic effect difficult. In this study, we compare the effect of four chemically characterized high molecular weight fucoidan extracts from Fucus distichus subsp. evanescens (FE_crude and fractions F1, F2, F3) on angiogenic and osteogenic processes in bone-related primary mono- and co-culture cell systems. By determining the gene expression and protein levels of the regulatory molecules vascular endothelial growth factor (VEGF), angiopoietin-1 (ANG-1), ANG-2 and stromal-derived factor 1 (SDF-1), we show that the extracted fucoidans negatively influence angiogenic and osteogenic processes in both the mono- and co-culture systems. We demonstrate that purer fucoidan extracts with a high fucose and sulfate content show stronger effects on these processes. Immunocytochemistry of the co-culture system revealed that treatment with FE_F3, containing the highest fucose and sulfate content, impaired the formation of angiogenic tube-like structures, indicating the anti-angiogenic properties of the tested fucoidans. This study highlights how chemical properties of fucoidan influence its bioactivity in a bone-related context and discusses how the observed phenotypes can be explained on a molecular level—knowledge that is indispensable for future therapies based on fucoidans.

Fucoidans, sulfated polysaccharides extracted from brown algae, are marine products with the potential to modulate bone formation and vascularization processes. The bioactivity and safety of fucoidans are highly associated with their chemical structure, which may vary with algae species and extraction method. Thus, in depth evaluation of fucoidan extracts in terms of endotoxin content, cytotoxicity, and their detailed molecular biological impact on the individual cell types in bone is needed. In this study, we characterized fucoidan extracts from three different Fucus species including Fucus vesiculosus (Fv), Fucus serratus (Fs), and Fucus distichus subsp. evanescens (Fe) for their chemical features, endotoxin content, cytotoxicity, and bioactive effects on human outgrowth endothelial cells (OEC) and human mesenchymal stem cells (MSC) as in vitro models for bone function and vascularization. Extracts contained mainly high molecular weight (HMW) fucoidans and were free of endotoxins that may cause inflammation or influence vascularization. OEC tolerated fucoidan concentrations up to 200 µg/mL, and no indication of cytotoxicity was observed. The inflammatory response, however, investigated by real-time PCR (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) and endothelial barrier assessed by impedance measurement differed for the individual extracts. MSC in comparison with endothelial cells were more sensitive to fucoidans and showed partly reduced metabolic activity and proliferation at higher doses of fucoidans. Further results for MSC indicated impaired osteogenic functions in alkaline phosphatase and calcification assays. All tested extracts consistently lowered important molecular mediators involved in angiogenesis, such a VEGF (vascular endothelial growth factor), ANG-1 (angiopoietin 1), and ANG-2 (angiopoietin 2), as indicated by RT-PCR and ELISA. This was associated with antiangiogenic effects at the functional level using selected extracts in co-culture models to mimic bone vascularization processes during bone regeneration or osteosarcoma.