Consensus guidelines for the use and interpretation of angiogenesis assays

The formation of new blood vessels, or angiogenesis, is a complex process that plays important roles in growth and development, tissue and organ regeneration, as well as numerous pathological conditions. Angiogenesis undergoes multiple discrete steps that can be individually evaluated and quantified by a large number of bioassays. These independent assessments hold advantages but also have limitations. This article describes in vivo, ex vivo, and in vitro bioassays that are available for the evaluation of angiogenesis and highlights critical aspects that are relevant for their execution and proper interpretation. As such, this collaborative work is the first edition of consensus guidelines on angiogenesis bioassays to serve for current and future reference

Modified Dental Composites for Bone Repair

The aim of this project was to develop modified dental composites for bone repair with additional remineralising monocalcium phosphate monohydrate (MCPM) and antibacterial polylysine (PLS). Bone composites were prepared with the powder to monomer weight ratio of 3:1. The monomer phase consisted of dimethacrylate monomers with chemical curing initiator and activator. Powder phase consisted of glass filler, PLS as antibacterial agent and MCPM as the mineralising agent. The level of remineralising agent and PLS were varied amongst formulations. Commercial polymethylmethacrylate (PMMA) (SimplexTM P) and bone composite (Cortoss®) were used for comparison. Degree of monomer conversion (DC) was assessed using fourier transform infrared spectroscopy (FTIR) (n=3). Biaxial flexural strength (BFS), tested by means of ball on ring technique, was used to compare the mechanical properties of each formulation (n=6). Surface apatite forming ability was carried out using scanning electron microscopy (SEM). Toxicity of monomers were assessed with sheep mesenchymal stem cells (MSCs) and MG63 cells using an Alamar Blue assay. Chick Embryo Chorioallantoic Membrane (CAM) assay was carried out to assess angiogenesis and development of a chick embryo upon contact with the experimental bone composite

Phosphatidylinositol 3-kinase/Akt signaling pathway and angiogenesis

Angiogenesis, the formation of new blood vessels from preexisting ones, is tightly controlled under physiological conditions, and deregulated angiogenesis contributes to many pathological situations. This study investigates the role of PI3K/Akt pathway in both physiological and pathological angiogenesis. Angiopoietin-1 (Ang1) is an endothelial specific growth factor that plays a critical role in vessel maturation and stabilization during angiogenesis. We found that Ang1 potently induced p70S6K1 activation in human umbilical vein endothelial cells (HUVECs). p70S6K1 is a downstream target of PI3K, but its role in angiogenesis has not be defined. In the work shown in Chapter 2, p70S6K1 activity in HUVECs was modified by adenovirus-mediated gene transfer, and we provided first evidence that p70S6K1 is directly involved in Ang1-induced angiogenic endothelial responses, including cell migration, invasion, survival, and capillary morphogenesis. We also examined the effect and mechanisms of action of insulin-like growth factor-I (IGF-I) on the expression of cyclooxygenase-2 (COX-2), which is a crucial player in angiogenesis and tumorigenesis. In Chapter 3, we showed that IGF-I efficiently upregulated COX-2 expression in human ovarian cancer cells, which was differentially regulated by PI3K, MAPK, and PKC signaling pathways at transcriptional and/or post-transcriptional levels. In the study shown in Chapter 4, we selectively modulated PI3K/Akt signaling in either human microvascular endothelial cells or cancer cells, and examined the effects on tumor angiogenesis using a chimeric tumor model. We found that PI3K and Akt activities in both endothelial cells and tumor cells contributed to tumor growth and angiogenesis, suggesting that targeting PI3K/Akt signaling in both cellular compartments may be more effective for anti-cancer therapy. In Chapter 5, we demonstrated that resveratrol has a strong inhibitory effects on hypoxiainducible factor 1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) expression in human ovarian cancer cells. The effects are associated with suppression of PI3K/Akt and MAPK pathways, interference with protein translational machinery, and enhancement of HIF-1alpha protein degradation through the proteasome. This work provides a better understanding of the molecular basis of angiogenesis, which we hope may facilitate the identification of novel therapeutic targets in the future