Chorioallantoic membrane assay as model for angiogenesis in tissue engineering : focus on stem cells

Tissue engineering aims to structurally and functionally regenerate damaged tissues, which requires the formation of new blood vessels that supply oxygen and nutrients by the process of angiogenesis. Stem cells are a promising tool in regenerative medicine due to their combined differentiation and paracrine angiogenic capacities. The study of their proangiogenic properties and associated potential for tissue regeneration requires complex in vivo models comprising all steps of the angiogenic process. The highly vascularized extraembryonic chorioallantoic membrane (CAM) of fertilized chicken eggs offers a simple, easy accessible, and cheap angiogenic screening tool compared to other animal models. Although the CAM assay was initially primarily performed for evaluation of tumor growth and metastasis, stem cell studies using this model are increasing. In this review, a detailed summary of angiogenic observations of different mesenchymal, cardiac, and endothelial stem cell types and derivatives in the CAM model is presented. Moreover, we focus on the variation in experimental setup, including the benefits and limitations of in ovo and ex ovo protocols, diverse biological and synthetic scaffolds, imaging techniques, and outcome measures of neovascularization. Finally, advantages and disadvantages of the CAM assay as a model for angiogenesis in tissue engineering in comparison with alternative in vivo animal models are described

Stem Cells for Cartilage Repair: Preclinical Studies and Insights in Translational Animal Models and Outcome Measures

Due to the restricted intrinsic capacity of resident chondrocytes to regenerate the lost cartilage postinjury, stem cell-based therapies have been proposed as a novel therapeutic approach for cartilage repair. Moreover, stem cell-based therapies using mesenchymal stem cells (MSCs) or induced pluripotent stem cells (iPSCs) have been used successfully in preclinical and clinical settings. Despite these promising reports, the exact mechanisms underlying stem cell-mediated cartilage repair remain uncertain. Stem cells can contribute to cartilage repair via chondrogenic differentiation, via immunomodulation, or by the production of paracrine factors and extracellular vesicles. But before novel cell-based therapies for cartilage repair can be introduced into the clinic, rigorous testing in preclinical animal models is required. Preclinical models used in regenerative cartilage studies include murine, lapine, caprine, ovine, porcine, canine, and equine models, each associated with its specific advantages and limitations. This review presents a summary of recent in vitro data and from in vivo preclinical studies justifying the use of MSCs and iPSCs in cartilage tissue engineering. Moreover, the advantages and disadvantages of utilizing small and large animals will be discussed, while also describing suitable outcome measures for evaluating cartilage repair

P availability and P leaching after reducing the mineral P fertilization and the use of digestate products as new organic fertilizers in a 4-year field trial with high P status

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Angiogenic Properties of 'Leukocyte- and Platelet-Rich Fibrin'

Leukocyte- and Platelet-Rich Fibrin (L-PRF) is an autologous platelet concentrate, consisting of a fibrin matrix enriched with platelets, leukocytes and a plethora of cytokines and growth factors. Since L-PRF is produced bedside from whole blood without the use of an anti-coagulant, it is becoming a popular adjuvant in regenerative medicine. While other types of platelet concentrates have been described to stimulate blood vessel formation, little is known about the angiogenic capacities of L-PRF. Therefore, this study aimed to fully characterize the angiogenic potential of L-PRF. With an antibody array, the growth factors released by L-PRF were determined and high levels of CXC chemokine receptor 2 (CXCR-2) ligands and epidermal growth factor (EGF) were found. L-PRF induced in vitro key steps of the angiogenic process: endothelial proliferation, migration and tube formation. In addition, we could clearly demonstrate that L-PRF is able to induce blood vessel formation in vivo, the chorioallantoic membrane assay. In conclusion, we could demonstrate the angiogenic capacity of L-PRF both in vitro and in vivo, underlying the clinical potential of this easy-to-use platelet concentrate.status: publishe

Preconditioning of Human Dental Pulp Stem Cells with Leukocyte- and Platelet-Rich Fibrin-Derived Factors Does Not Enhance Their Neuroregenerative Effect

Pathologies of the central nervous system are characterized by loss of brain tissue and neuronal function which cannot be adequately restored by endogenous repair processes. This stresses the need for novel treatment options such as cell-based therapies that are able to restore damaged tissue or stimulate repair. This study investigated the neuroregenerative potential of the conditioned medium of human dental pulp stem cells (CM-hDPSCs) on neural stem cell (NSC) proliferation and migration as well as on neurite outgrowth of primary cortical neurons (pCNs). Additionally, the effect of leukocyte- and platelet-rich fibrin (L-PRF) priming on the neuroregenerative potential of the hDPSC secretome on NSCs and pCNs was evaluated. L-PRF contains factors that enhance stem cell-induced regeneration, but its effect on hDPSC-mediated neuroregeneration is unknown. This study demonstrated that CM-hDPSCs enhanced neuritogenesis. Moreover, CM-hDPSCs had a chemoattractant effect on NSCs. Although priming hDPSCs with L-PRF increased brain-derived neurotrophic factor secretion, no additional effects on the paracrine-mediated repair mechanisms were observed. These data support the neuroregenerative potential of hDPSCs, and although priming had no additional effect, the potential of L-PRF-primed hDPSCs on distinct regenerative mechanisms remains to be clarified

Evidence-based sustainable phosphorus use in agriculture in Flanders (Belgium)

Phosphorus (P) is an important nutrient for agriculture but excess P application to soils can contribute to eutrophication of surface waters. Phosphorus fertilisation recommendations rarely take environmental concerns into account (Jordan-Meille et al., 2012). Soil P tests used for soil P content measurements mostly build on chemical extractions for which empirical (not mechanistic) relations with crop responses are determined. Recently, the Flemish Land Agency (government of Flanders, Belgium) initiated a research project to stimulate sustainable phosphorus use in agriculture. Three research institutes (Institute for Agricultural and Fisheries Research (ILVO), Soil Service of Belgium and KU Leuven) started this four-year project at the beginning of 2015. In the first phase of the project, soil P tests are evaluated in order to select the (combination of) soil P test(s) that best reflect both (i) the P availability for plants and (ii) the risk of P losses towards surface waters at a relatively low cost. Several common and new tests (ammonium lactate extraction, 0.01 M CaCl2 extraction, oxalate extraction, Olsen extraction, etc) are compared in a pot experiment in a depletion scenario and on soil samples from 14 long term fertilisation field trials in NW Europe with reported yield differences due to soil P differences. Suitability of the test is assessed by the correlation with crop yield and the relative width of the 95% confidence interval of the critical soil P content, i.e. the soil P content corresponding with 95% relative yield. Not only single soil P tests but also combinations are evaluated. According to van Rotterdam-Los (2010), especially the combination of a soil P test reflecting P intensity (directly available P) and one reflecting P quantity (P available in the long term) shows promise for describing the behaviour and availability of P. The same tests are also evaluated for correlations with soil P losses by performing soil column leaching experiments under unsaturated conditions. The selected test(s) will be used in the second and third parts of the research project. In this second part, the soil P contents at which yields are optimal and losses still small, will be defined as the target zone. In the third part, we will derive soil P fertilisation advice in order to reach or remain in the target zone from sorption/desorption experiments and field trials. The outline of this project and its first results for sustainable P use will be presented and discussed.status: publishe